Compositions and methods for combination antiviral therapy

ABSTRACT

The present invention relates to therapeutic combinations of [2-(6-amino-purin-9-yl)-1-methyl-ethoxymethyl]-phosphonic acid diisopropoxycarbonyloxymethyl ester (tenofovir disoproxil fumarate, Viread®) and (2R,5S,cis)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-pyrimidin-2-one (emtricitabine, Emtriva™, (−)-cis FTC) and their physiologically functional derivatives. The combinations may be useful in the treatment of HIV infections, including infections with HIV mutants bearing resistance to nucleoside and/or non-nucleoside inhibitors. The present invention is also concerned with pharmaceutical compositions and formulations of said combinations of tenofovir disoproxil fumarate and emtricitabine, and their physiologically functional derivatives, as well as therapeutic methods of use of those compositions and formulations.

This non-provisional application is a continuation of U.S. patentapplication Ser. No. 10/540,794, filed Mar. 20, 2006, now abandonedwhich is a national stage entry of PCT/US04/00832, filed Jan. 13, 2004which claims the benefit of Provisional Application Nos. 60/440,246 and60/440,308, both filed Jan. 14, 2003, which is incorporated herein byreference.

FIELD OF THE INVENTION

The invention relates generally to combinations of compounds withantiviral activity and more specifically with anti-HIV properties. Inparticular, it relates to chemically stable combinations of structurallydiverse anti-viral agents.

BACKGROUND OF THE INVENTION

Human immunodeficiency virus (HIV) infection and related diseases are amajor public health problem worldwide. Human immunodeficiency virus type1 (HIV-1) encodes at least three enzymes which are required for viralreplication: reverse transcriptase (RT), protease (Prt), and integrase(Int). Although drugs targeting reverse transcriptase and protease arein wide use and have shown effectiveness, particularly when employed incombination, toxicity and development of resistant strains have limitedtheir usefulness (Palella, et al N. Engl. J. Med. (1998) 338:853-860;Richman, D. D. Nature (2001) 410:995-1001). Human immunodeficiency virustype 1 (HIV-1) protease (Prt) is essential for viral replication and isan effective target for approved antiviral drugs. The HIV Pit cleavesthe viral Gag and Gag-Pol polyproteins to produce viral structuralproteins (p17, p24, p7 and p6) and the three viral enzymes. Combinationtherapy with RT inhibitors has proven to be highly effective insuppressing viral replication to unquantifiable levels for a sustainedperiod of time. Also, combination therapy with RT and Prt inhibitors(PI) have shown synergistic effects in suppressing HIV replication.Unfortunately, a high percentage, typically 30 to 50% of patientscurrently fail combination therapy due to the development of drugresistance, non-compliance with complicated dosing regimens,pharmacokinetic interactions, toxicity, and lack of potency. Therefore,there is a need for new HIV-1 inhibitors that are active against mutantHIV strains, have distinct resistance profiles, fewer side effects, lesscomplicated dosing schedules, and are orally active. In particular,there is a need for a less onerous dosage regimen, such as once per dayoral dosing, optimally with as few pills as possible.

The use of combinations of compounds can yield an equivalent antiviraleffect with reduced toxicity, or an increase in drug efficacy. Loweroverall drug doses can reduce the frequency of occurrence ofdrug-resistant variants of HIV. Many different methods have been used toexamine the effects of combinations of compounds acting together indifferent assay systems (Furman WO 02/068058). Lower doses predictbetter patient compliance when pill burden decreases, dosing schedulesare simplified and, optionally, if synergy between compounds occurs(Loveday, C. “Nucleoside reverse transcriptase inhibitor resistance”(2001) JAIDS Journal of Acquired Immune Deficiency Syndromes26:S10-S24). AZT (Zidovudine™, 3′,-azido, 3′-deoxythymidine)demonstrates synergistic antiviral activity in vitro in combination withagents that act at HIV-1 replicative steps other than reversetranscription, including recombinant soluble CD4 castanospermine andrecombinant interferon-α. However, it must be noted that combinations ofcompounds can give rise to increased cytotoxicity. For example, AZT andrecombinant interferon-α have an increased cytotoxic effect on normalhuman bone marrow progenitor cells.

Chemical stability of combinations of antiviral agents is an importantaspect of co-formulation success and the present invention providesexamples of such combinations.

There is a need for new combinations of orally-active drugs for thetreatment of patients infected with certain viruses, e.g. HIV, thatprovide enhanced therapeutic safety and efficacy, impart lowerresistance, and predict higher patient compliance.

SUMMARY OF THE INVENTION

The present invention provides combinations of antiviral compounds, inparticular compositions and methods for inhibition of HIV. In anexemplary aspect, the invention includes a composition includingtenofovir disoproxil fumarate and emtricitabine which has anti-HIVactivity. The composition of tenofovir DF and emtricitabine is bothchemically stable and either synergistic and/or reduces the side effectsof one or both of tenofovir DF and emtricitabine. Increased patientcompliance is likely in view of the lower pill burden and simplifieddosing schedule.

The present invention relates to therapeutic combinations of[2-(6-amino-purin-9-yl)-1-methyl-ethoxymethyl]-phosphonic aciddiisopropoxycarbonyloxymethyl ester fumarate (tenofovir disoproxilfumarate, tenofovir DF, TDF, Viread®) and(2R,5S,cis)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-pyrimidin-2-one(emtricitabine, Emtriva™, (−)-cis FTC) and their use in the treatment ofHIV infections including infections with HIV mutants bearing resistanceto nucleoside and/or non-nucleoside inhibitors. The present invention isalso concerned with pharmaceutical compositions and formulations of saidcombinations of tenofovir disoproxil fumarate and emtricitabine. Anotheraspect of the invention is a pharmaceutical formulation comprising aphysiologically functional derivative of tenofovir disoproxil fumarateor a physiologically functional derivative of emtricitabine.

Therapeutic combinations and pharmaceutical compositions andformulations of the invention include the combination of PMEA or PMPA(tenofovir) compounds with emtricitabine or(2R,5S,cis)-4-amino-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-pyrimidin-2-one(3TC, lamivudine, Epivir™), and their use in the treatment of HIVinfections.

One aspect of the invention is a method for the treatment or preventionof the symptoms or effects of an HIV infection in an infected animalwhich comprises administering to, i.e. treating, said animal with atherapeutically effective amount of a combination comprising[2-(6-amino-purin-9-yl)-1-methyl-ethoxymethyl]-phosphonic aciddiisopropoxycarbonyloxymethyl ester fumarate (tenofovir DF, TDF) or aphysiologically functional derivative thereof and(2R,5S,cis)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-pyrimidin-2-one(emtricitabine) or a physiologically functional derivative thereof.

Another aspect of the invention is a unit dosage form of a therapeuticcombination comprising tenofovir disoproxil fumarate and emtricitabine,or physiological functional derivatives thereof. The unit dosage formmay be formulated for administration by oral or other routes and isunexpectedly chemically stable in view of the properties of thestructurally diverse components.

Another aspect of the invention is directed to chemically stablecombination antiviral compositions comprising tenofovir disoproxilfumarate and emtricitabine. In a further aspect of the invention, thechemically stable combinations of tenofovir disoproxil fumarate andemtricitabine further comprise a third antiviral agent. In thisthree-component mixture, the unique chemical stability of tenofovirdisoproxil fumarate and emtricitabine is taken advantage of in order toenable the combination with the third antiviral agent. Particularlyuseful third agents include, by way of example and not limitation, thoseof Table A. Preferably, the third component is an agent approved forantiviral use in humans, more preferably, it is an NNRTI or a proteaseinhibitor (PI), more preferably yet, it is an NNRTI. In a particularlypreferred embodiment, the invention is directed to a combination of thechemically stable mixture of tenofovir disoproxil fumarate andemtricitabine together with efavirenz.

Another aspect of the invention is a patient pack comprising at leastone, typically two, and optionally, three active ingredients and otherantiviral agents selected from tenofovir disoproxil fumarate andemtricitabine, and an information insert containing directions on theuse of tenofovir disoproxil fumarate and emtricitabine together incombination.

Another aspect of the invention is a process for preparing thecombinations hereinbefore described, which comprises bringing intoassociation tenofovir DF and emtricitabine of the combination in amedicament to provide an antiviral effect. In a further aspect of thepresent invention, there is provided the use of a combination of thepresent invention in the manufacture of a medicament for the treatmentof any of the aforementioned viral infections or conditions.

DETAILED DESCRIPTION OF THE INVENTION

While the invention will be described in conjunction with the enumeratedclaims, it will be understood that they are not intended to limit theinvention to those claims. On the contrary, the invention is intended tocover all alternatives, modifications, and equivalents, which may beincluded within the scope of the present invention as defined by theclaims.

DEFINITIONS

Unless stated otherwise, the following terms and phrases as used hereinare intended to have the following meanings:

When tradenames are used herein, applicants intend to independentlyinclude the tradename product and the active pharmaceuticalingredient(s) of the tradename product.

The term “chemical stability” means that the two primary antiviralagents in combination are substantially stable to chemical degradation.Preferably, they are sufficiently stable in physical combination topermit commercially useful shelf life of the combination product.Typically, “chemically stable” means that a first component of themixture does not act to degrade a second component when the two arebrought into physical combination to form a pharmaceutical dosage form.More typically, “chemically stable” means that the acidity of a firstcomponent does not catalyzes or otherwise accelerate the aciddecomposition of a second component. By way of example and notlimitation, in one aspect of the invention, “chemically stable” meansthat tenofovir disoproxil fumarate is not substantially degraded by theacidity of emtricitabine. “Substantially” in this context means at leastabout less than 10%, preferably less than 1%, more preferably less than0.1%, more preferably yet, less than 0.01% acid degradation of tenofovirdisoproxil fumarate over a 24-hour period when the products are in apharmaceutical dosage form.

The terms “synergy” and “synergistic” mean that the effect achieved withthe compounds used together is greater than the sum of the effects thatresults from using the compounds separately, i.e. greater than whatwould be predicted based on the two active ingredients administeredseparately. A synergistic effect may be attained when the compounds are:(1) co-formulated and administered or delivered simultaneously in acombined formulation; (2) delivered by alternation or in parallel asseparate formulations; or (3) by some other regimen. When delivered inalternation therapy, a synergistic effect may be attained when thecompounds are administered or delivered sequentially, e.g. in separatetablets, pills or capsules, or by different injections in separatesyringes. In general, during alternation therapy, an effective dosage ofeach active ingredient is administered sequentially, i.e. serially,whereas in combination therapy, effective dosages of two or more activeingredients are administered together. A synergistic antiviral effectdenotes an antiviral effect which is greater than the predicted purelyadditive effects of the individual compounds of the combination.

The term “physiologically functional derivative” means apharmaceutically active compound with equivalent or near equivalentphysiological functionality to tenofovir DF or emtricitabine whenadministered in combination with another pharmaceutically activecompound in a combination of the invention. As used herein, the term“physiologically functional derivative” includes any: physiologicallyacceptable salt, ether, ester, prodrug, solvate, stereoisomer includingenantiomer, diastereomer or stereoisomerically enriched or racemicmixture, and any other compound which upon administration to therecipient is capable of providing (directly or indirectly) such acompound or an antivirally active metabolite or residue thereof.

“Bioavailability” is the degree to which the pharmaceutically activeagent becomes available to the target tissue after the agent'sintroduction into the body. Enhancement of the bioavailability of apharmaceutically active agent can provide a more efficient and effectivetreatment for patients because, for a given dose, more of thepharmaceutically active agent will be available at the targeted tissuesites.

The compounds of the combinations of the invention may be referred to as“active ingredients” or “pharmaceutically active agents.”

The term “prodrug” as used herein refers to any compound that whenadministered to a biological system generates the drug substance, i.e.active ingredient, as a result of spontaneous chemical reaction(s),enzyme catalyzed chemical reaction(s), and/or metabolic chemicalreaction(s).

“Prodrug moiety” means a labile functional group which separates fromthe active inhibitory compound during metabolism, systemically, inside acell, by hydrolysis, enzymatic cleavage, or by some other process(Bundgaard, Hans, “Design and Application of Prodrugs” in Textbook ofDrug Design and Development (1991), P. Krogsgaard-Larsen and H.Bundgaard, Eds. Harwood Academic Publishers, pp. 113-191). Prodrugmoieties can serve to enhance solubility, absorption and lipophilicityto optimize drug delivery, bioavailability and efficacy. A “prodrug” isthus a covalently modified analog of a therapeutically-active compound.

“Alkyl” means a saturated or unsaturated, branched, straight-chain,branched, or cyclic hydrocarbon radical derived by the removal of onehydrogen atom from a single carbon atom of a parent alkane, alkene, oralkyne. Typical alkyl groups consist of 1-18 saturated and/orunsaturated carbons, such as normal, secondary, tertiary or cycliccarbon atoms. Examples include, but are not limited to: methyl, Me(—CH₃), ethyl, Et (—CH₂CH₃), acetylenic (—C≡CH), ethylene, vinyl(—CH═CH₂), 1-propyl, n-Pr, n-propyl (—CH₂CH₂CH₃), 2-propyl, i-Pr,i-propyl (—CH(CH₃)₂), allyl (—CR₂CH═CH₂), propargyl (—CH₂C≡CH),cyclopropyl (—C₃H₅), 1-butyl, n-Bu, n-butyl (—CH₂CH₂CH₂CH₃),2-methyl-1-propyl, i-Bu, 1-butyl (—CH₂CH(CH₃)₂), 2-butyl, s-Bu, s-butyl(—CH(CH₃)CH²CH₃), 2-methyl-2-propyl, t-Bu, t-butyl (—C(CH₃)₃), 1-pentyl,n-pentyl, (—CH₂CH₂CH₂CH₂CH₃), 2-pentyl (—CH(CH₃)CH₂CH₂CH₃), 3-pentyl(—CH(CH₂CH₃)₂), 2-methyl-2-butyl (—C(CH₃)₂CH₂CH₃), cyclopentyl (—C₅H₉),3-methyl-2-butyl (—CH(CH₃)CH(CH₃)₂), 3-methyl-1-butyl (—CH₂CH₂CH(CH₃)₂),2-methyl-1-butyl (—CH₂CH(CH₃)CH₂CH₃), 1-hexyl (—CH₂CH₂CH₂CH₂CH₂CH₃),5-hexenyl (—CH₂CH₂CH₂CH₂CH═CH₂) 1-hexyl (—CH(CH₃)CH₂CH₂CH₂CH₃), 3-hexyl(—CH(CH₂CH₃)(CH₂CH₂CH₃)), cyclohexyl (—C₆H₁₁), 2-methyl-2-pentyl(—C(CH₃)₂CH₂CH₂CH₃), 3-methyl-2-pentyl (—CH(CH₃)CH(CH₃)CH₂CH₃),4-methyl-2-pentyl (—CH(CH₃)CH₂CH(CH₃)₂), 3-methyl-3-pentyl(—C(CH₃)(CH₂CH₃)₂), 2-methyl-3-pentyl (—CH(CH₂CH₃)CH(CH₃)₂),2,3-dimethyl-2-butyl (—C(CH₃)₂CH(CH₃)₂), and 3,3-dimethyl-2-butyl(—CH(CH₃)C(CH₃)₃.

“Aryl” means a monovalent aromatic hydrocarbon radical of 6-20 carbonatoms derived by the removal of one hydrogen atom from a single carbonatom of a parent aromatic ring system. Typical aryl groups include, butare not limited to, radicals derived from benzene, substituted benzene,naphthalene, anthracene, biphenyl, and the like.

“Arylalkyl” refers to an acyclic alkyl radical in which one of thehydrogen atoms bonded to a carbon atom, typically a terminal or Sp³carbon atom, is replaced with an aryl radical. Typical arylalkyl groupsinclude, but are not limited to, benzyl, 2-phenylethan-1-yl,2-phenylethen-1-yl, naphthylmethyl, 2-naphthylethan-1-yl,2-naphthylethen-1-yl, naphthobenzyl, 2-naphthophenylethan-1-yl and thelike. The arylalkyl group 6 to 20 carbon atoms e.g., the alkyl moiety,including alkanyl, alkenyl or alkynyl groups, of the arylalkyl group is1 to 6 carbon atoms and the aryl moiety is 5 to 14 carbon atoms.

“Substituted alkyl”, “substituted aryl”, and “substituted arylalkyl”mean alkyl, aryl, and arylalkyl respectively, in which one or morehydrogen atoms are each independently replaced with a substituent.Typical substituents include, but are not limited to, —X, —R, —O⁻, —OR,—SR, —S⁻, —NR₂, —NR₃, ═NR, —CX₃, —CN, —OCN, —SCN, —N═C—O, —NCS, —NO,—NO₂, ═N₂, —N₃, NC(═O)R, —C(═O)R, —C(═O)NRR—S(═O)₂O⁻, —S(═O)₂OH,—S(═O)₂R, —OS(═O)OR, —S(═O)₂NR, —S(═O)R, —OP(═O)O₂RR, —P(═O)O₂RR—P(═O)(O⁻)₂, —P(═O)(OH)₂, —C(═O)R, —C(═O)X, —C(S)R, —C(O)OR, —C(O)O⁻,—C(S)OR, —C(O)SR, —C(S)SR, —C(O)NRR, —C(S)NRR, —C(NR)NRR, where each Xis independently a halogen: F, Cl, Br, or I; and each R is independently—H, alkyl, aryl, heterocycle, or prodrug moiety.

“Heteroaryl” and “Heterocycle” refer to a ring system in which one ormore ring atoms is a heteroatom, e.g. nitrogen, oxygen, and sulfur.Heterocycles are described in: Katritzky, Alan R., Rees, C. W., andScriven, E. Comprehensive Heterocyclic Chemistry (1996) Pergamon Press;Paquette, Leo A.; Principles of Modern Heterocyclic Chemistry W. A.Benjamin, New York, (1968), particularly Chapters 1, 3, 4, 6, 7, and 9;“The Chemistry of Heterocyclic Compounds, A series of Monographs” (JohnWiley & Sons, New York, 1950 to present), in particular Volumes 13, 14,16, 19, and 28. Exemplary heterocycles include but are not limited tosubstituents, i.e. radicals, derived from pyrrole, indole, furan,benzofuran, thiophene, benzothiophene, 2-pyridyl, 3-pyridyl, 4-pyridyl,2-quinolyl, 3-quinolyl, 4-quinolyl, 2-imidazole, 4-imidazole,3-pyrazole, 4-pyrazole, pyridazine, pyrimidine, pyrazine, purine,cinnoline, pthalazine, quinazoline, quinoxaline, 3-(1,2,4-N)-triazolyl,5-(1,2,4-N)-triazolyl, 5-tetrazolyl, 4-(1-O,3-N)-oxazole,5-(1-O,3-N)-oxazole, 4-(1-S,3-N)-thiazole, 5-(1-S,3-N)-thiazole,2-benzoxazole, 2-benzothiazole, 4-(1,2,3-N)-benzotriazole, andbenzimidazole.

Stereochemical definitions and conventions used herein generally followS. P. Parker, Ed., McGraw-Hill Dictionary of Chemical Terms (1984)McGraw-Hill Book Company, New York; and Eliel, E. and Wilen, S.,Stereochemistry of Organic Compounds (1994) John Wiley & Sons, Inc., NewYork. Many organic compounds exist in optically active forms, i.e., theyhave the ability to rotate the plane of plane-polarized light. Indescribing an optically active compound, the prefixes D and L or R and Sare used to denote the absolute configuration of the molecule about itschiral center(s). The prefixes d and l or (+) and (−) are employed todesignate the sign of rotation of plane-polarized light by the compound,with (−) or 1 meaning that the compound is levorotatory. A compoundprefixed with (+) or d is dextrorotatory. For a given chemicalstructure, these compounds, called stereoisomers, are identical exceptthat they are mirror images of one another. A specific stereoisomer isalso referred to as an enantiomer, and a mixture of such isomers isoften called an enantiomeric mixture. A 50:50 mixture of enantiomers isreferred to as a racemic mixture or a racemate. The terms “racemicmixture” and “racemate” refer to an equimolar mixture of twoenantiomeric species, devoid of optical activity.

The term “chiral” refers to molecules which have the property ofnon-superimposability of the mirror image partner, while the term“achiral” refers to molecules which are superimposable on their mirrorimage partner.

The term “stereoisomers” refers to compounds which have identicalchemical constitution, but differ with regard to the arrangement of theatoms or groups in space.

“Diastereomer” refers to a stereoisomer with two or more centers ofchirality and whose molecules are not mirror images of one another.Diastereomers have different physical properties, e.g. melting points,boiling points, spectral properties, and reactivities. Mixtures ofdiastereomers may separate under high resolution analytical proceduressuch as electrophoresis and chromatography.

“Enantiomers” refer to two stereoisomers of a compound which arenon-superimposable mirror images of one another.

Active Ingredients of the Combinations

The present invention provides novel combinations of two or more activeingredients being employed together. In some embodiments, a synergisticantiviral effect is achieved. In other embodiments, a chemically stablecombination is obtained. The combinations include at least one activeingredient selected from (1) tenofovir disoproxil fumarate andphysiologically functional derivatives, and at least one activeingredient selected from (2) emtricitabine and physiologicallyfunctional derivatives. The term “synergistic antiviral effect” is usedherein to denote an antiviral effect which is greater than the predictedpurely additive effects of the individual components (a) and (b) of thecombination.

Tenofovir disoproxil fumarate (also known as Viread®, Tenofovir DF,Tenofovir disoproxil, TDF, Bis-POC-PMPA (U.S. Pat. Nos. 5,935,946,5,922,695, 5,977,089, 6,043,230, 6,069,249) is a prodrug of tenofovir,and has the structure:

and including fumarate salt (HO₂CCH₂CH₂CO₂ ⁻).

The chemical names for Tenofovir disoproxil include:[2-(6-amino-purin-9-yl)-1-methyl-ethoxymethyl]-phosphonic aciddiisopropoxycarbonyloxymethyl ester;9-[(R)-2-[[bis[[(isopropoxycarbonyl)oxy]methoxy]phosphinyl]methoxy]propyl]adenine;and 2,4,6,8-tetraoxa-5-phosphanonanedioic acid,5-[[(1R)-2-(6-amino-9H-purin-9-yl)-1-methylethoxy]methyl]-,bis(1-methylethyl)ester, 5-oxide. The CAS Registry numbers include:201341-05-1; 202138-50-9; 206184-49-8. It should be noted that theethoxymethyl unit of tenofovir has a chiral center. The R (rectus, righthanded configuration) enantiomer is shown. However, the invention alsoincludes the S isomer. The invention includes all enantiomers,diastereomers, racemates, and enriched stereoisomer mixtures oftenofovir (PMPA) and physiologically functional derivatives thereof.

PMPA or tenofovir (U.S. Pat. Nos. 4,808,716, 5,733,788, 6,057,305) hasthe structure:

The chemical names of PMPA, tenofovir include:(R)-9-(2-phosphonylmethoxypropyl)adenine; and phosphonic acid,[[(1R)-2-(6-amino-9H-purin-9-yl)-1-methylethoxy]methyl]. The CASRegistry number is 147127-20-6.

Tenofovir disoproxil fumarate (DF) is a nucleotide reverse transcriptaseinhibitor approved in the United States in 2001 for the treatment ofHIV-1 infection in combination with other antiretroviral agents.Tenofovir disoproxil fumarate or Viread® (Gilead Science, Inc.) is thefumarate salt of tenofovir disoproxil. Viread® may be named as: 9-[;R)-2-[[bis[[(isopropoxycarbonyl)oxy]methoxy]phosphinyl]methoxy]propyl]adeninefumarate (1:1); or 2,4,6,8-tetraoxa-5-phosphanonanedioic acid,5-[[(1R)-2-(6-amino-9H-purin-9-yl)-1-methylethoxy]methyl]-,bis(1-methylethyl) ester, 5-oxide, (2E)-2-butenedioate (1:1). The CASRegistry number is 202138-50-9.

Physiologically functional derivatives of tenofovir disoproxil fumarateinclude PMEA (adefovir, 9-((R)-2-(phosphonomethoxy)ethyl)adenine) andPMPA compounds. Exemplary combinations include a PMEA or PMPA compoundin combination with emtricitabine or 3TC. PMEA and PMPA compounds havethe structures:

where PMEA (R³ is H) and PMPA (R³ is C₁-C₆ alkyl, C₁-C₆ substitutedalkyl, or CH₂OR⁸ where R⁸ is C₁-C₆ alkyl, C₁-C₆ hydroxyalkyl or C₁-C₆haloalkyl. R⁶ and R⁷ are independently H or C₁-C₆ alkyl. R⁴ and R⁵ areindependently H, NH₂, NHR or NR₂ where R is C₁-C₆ alkyl. R¹ and R² areindependently H, C₁-C₆ alkyl, C₁-C₆ substituted alkyl, C₆-C₂₀ aryl,C₆-C₂₀ substituted aryl, C₆-C₂₀ arylalkyl, C₆-C₂₀ substituted arylalkyl,acyloxymethyl esters —CH₂C(═O)R⁹ (e.g. POM) or acyloxymethyl carbonates—CH₂OC(═O)OR⁹ (e.g. POC) where R⁹ is C₁-C₆ alkyl, C₁-C₆ substitutedalkyl, C₆-C₂₀ aryl or C₆-C₂₀ substituted aryl. For example, R₁ and R₂may be pivaloyloxymethoxy, POM, —CH₂C(═O)C(CH₃)₃; CH₂OC(═O)OC(CH₃)₃; orPOC, —CH₂C(═O)OCH(CH₃)₂. Also for example, tenofovir has the structurewhere R³ is CH₃, and R¹, R², R⁴, R⁵, R⁶ and R⁷ are H. Dialkylphosphonates may be prepared according to the methods of: Quast et al(1974) Synthesis 490; Stowell et al (1990) Tetrahedron Lett. 3261; U.S.Pat. No. 5,663,159.

The PMPA compound may be enantiomerically-enriched or purified (singlestereoisomer) where the carbon atom bearing R³ may be the R or Senantiomer. The PMPA compound may be a racemate, i.e. a mixture of R andS stereoisomers.

Adefovir (9-(2-phosphonomethoxyethyl)adenine where R₁-R₇═H) is anexemplary PMEA compound (U.S. Pat. Nos. 4,808,716, 4,724,233). As thebis-pivalate prodrug, Adefovir dipivoxil, also known as bis-POM PMEA,(R₃-R₇═H, R₁ and R₂=—CH₂CO(═O)C(CH₃)₃, pivoxil, POM,pivaloyloxymethoxy), is effective against HIV and Hepatitis B infections(U.S. Pat. Nos. 5,663,159, 6,451,340). Adefovir dipivoxil hasdemonstrated minor to moderate synergistic inhibition of HIV replicationin combination with other compounds with anti-HIV activity includingPMPA, d4T, ddC, nelfinavir, ritonavir, and saquinavir (Mulato et al(1997) Antiviral Research 36:91-97).

The invention includes all enantiomers, diastereomers, racemates, andenriched stereoisomer mixtures of PMEA and PMPA, and physiologicallyfunctional derivatives thereof.

Emtricitabine ((−)-cis-FTC, Emtriva™), a single enantiomer of FTC, is apotent nucleoside reverse transcriptase inhibitor approved for thetreatment of HIV (U.S. Pat. Nos. 5,047,407, 5,179,104, 5,204,466,5,210,085, 5,486,520, 5,538,975, 5,587,480, 5,618,820, 5,763,606,5,814,639, 5,914,331, 6,114,343, 6,180,639, 6,215,004; WO 02/070518).The single enantiomer emtricitabine has the structure:

The chemical names for emtricitabine include: (−)-cis-FTC;β-L-hydroxymethyl-5-(5-fluorocytosin-1-yl)-1,3-oxathiolane;(2R,5S)-5-fluoro-1-[2-(hydroxymethyl)-1,3-oxathiolan-5-yl]cytosine; and4-amino-5-fluoro-1-(2-hydroxymethyl-[1,3]-(2R,5S)-oxathiolan-5-yl)-1H-pyrimidin-2-one.The CAS Registry numbers include: 143491-57-0; 143491-54-7. It should benoted that FTC contains two chiral centers, at the 2 and 5 positions ofthe oxathiolane ring, and therefore can exist in the form of two pairsof optical isomers (i.e. enantiomers) and mixtures thereof includingracemic mixtures. Thus, FTC may be either a cis or a trans isomer ormixtures thereof. Mixtures of cis and trans isomers are diastereomerswith different physical properties. Each cis and trans isomer can existas one of two enantiomers or mixtures thereof including racemicmixtures. The invention includes all enantiomers, diastereomers,racemates, and enriched stereoisomer mixtures of emtricitabine andphysiologically functional derivatives thereof. For example, theinvention includes physiological functional derivatives such as the 1:1racemic mixture of the enantiomers(2R,5S,cis)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-pyrimidin-2-one(emtricitabine) and its mirror image (2S,5R,cis)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-pyrimidin-2-one,or mixtures of the two enantiomers in any relative amount. The inventionalso includes mixtures of cis and trans forms of FTC.

Physiologically functional derivatives of emtricitabine include 1,3oxathiolane nucleosides having the structure:

In the 1,3 oxathiolane nucleoside structure above, B is a nucleobaseincluding any nitrogen-containing heterocyclic moiety capable of formingWatson-Crick hydrogen bonds in pairing with a complementary nucleobaseor nucleobase analog, e.g. a purine, a 7-deazapurine, or a pyrimidine.Examples of B include the naturally occurring nucleobases: adenine,guanine, cytosine, uracil, thymine, and minor constituents and analogsof the naturally occurring nucleobases, e.g. 7-deazaadenine,7-deazaguanine, 7-deaza-8-azaguanine, 7-deaza-8-azaadenine, inosine,nebularine, nitropyrrole, nitroindole, 2-aminopurine,2-amino-6-chloropurine, 2,6-diaminopurine, hypoxanthine, pseudouridine,5-fluorocytosine, 5-chlorocytosine, 5-bromocytosine, 5-iodocytosine,pseudocytosine, pseudoisocytosine, 5-propynylcytosine, isocytosine,isoguanine, 7-deazaguanine, 2-thiopyrimidine, 6-thioguanine,4-thiothymine, 4-thiouracil, O⁶-methylguanine, N⁶-methyladenine,O⁴-methylthymine, 5,6-dihydrothymine, 5,6-dihydrouracil, 4-methylindole,pyrazolo[3,4-D]pyrimidines (U.S. Pat. Nos. 6,143,877 and 6,127,121; WO01/38584), and ethenoadenine (Fasman (1989) in Practical Handbook ofBiochemistry and Molecular Biology, pp. 385-394, CRC Press, Boca Raton,Fla.).

Nucleobases B may be attached in the configurations ofnaturally-occurring nucleic acids to the 1,3 oxathiolane moiety througha covalent bond between the N-9 of purines, e.g. adenin-9-yl andguanin-9-yl, or N-1 of pyrimidines, e.g. thymin-1-yl and cytosin-1-yl(Blackburn, G. and Gait, M. Eds. “DNA and RNA structure” in NucleicAcids in Chemistry and Biology, 2^(nd) Edition, (1996) Oxford UniversityPress, pp. 15-81).

Also in the 1,3 oxathiolane nucleoside structure above, R is H, C₁-C₁₈alkyl, C₁-C₁₈ substituted alkyl, C₂-C₁₈ alkenyl, C₂-C₁₈ substitutedalkenyl, C₂-C₁₈ alkynyl, C₂-C₁₈ substituted alkynyl, C₆-C₂₀ aryl, C₆-C₂₀substituted aryl, C₂-C₂₀ heterocycle, C₂-C₂₀ substituted heterocycle,phosphonate, phosphophosphonate, diphosphophosphonate, phosphate,diphosphate, triphosphate, polyethyleneoxy, or a prodrug moiety.

Physiologically functional derivatives of emtricitabine also include 3TC(lamivudine, Epivir®), a reverse transcriptase inhibitor approved in theUnited States for the treatment of HIV-1 infection in combination withAZT as Combivir® (GlaxoSmithKline). U.S. Pat. Nos. 5,859,021; 5,905,082;6,177,435; 5,627,186; 6,417,191. Lamivudine (U.S. Pat. Nos. 5,587,480,5,696,254, 5,618,820, 5,756,706, 5,744,596, 568,164, 5,466,806,5,151,426) has the structure:

For example and for some therapeutic uses, 3TC may be a physiologicallyfunctional derivative of emtricitabine in combination with tenofovir DFor a physiologically functional derivative of tenofovir DF.

It will be appreciated that tenofovir DF and emtricitabine, and theirphysiologically functional derivatives may exist in keto or enoltautomeric forms and the use of any tautomeric form thereof is withinthe scope of this invention. Tenofovir DF and emtricitabine willnormally be utilized in the combinations of the invention substantiallyfree of the corresponding enantiomer, that is to say no more than about5% w/w of the corresponding enantiomer will be present.

Prodrugs

The invention includes all prodrugs of tenofovir and emtricitabine. Anexemplary prodrug of tenofovir is tenofovir disoproxil fumarate (TDF,Viread®). A large number of structurally-diverse prodrugs have beendescribed for phosphonic acids (Freeman and Ross in Progress inMedicinal Chemistry 34: 112-147 (1997). A commonly used prodrug class isthe acyloxyalkyl ester, which was first used as a prodrug strategy forcarboxylic acids and then applied to phosphates and phosphonates byFarquhar et al (1983) J. Pharm. Sci. 72: 324; also U.S. Pat. Nos.4,816,570, 4,968,788, 5,663,159 and 5,792,756. Subsequently, theacyloxyalkyl ester was used to deliver phosphonic acids across cellmembranes and to enhance oral bioavailability. A close variant of theacyloxyalkyl ester strategy, the alkoxycarbonyloxyalkyl ester, may alsoenhance oral bioavailability as a prodrug moiety in the compounds of thecombinations of the invention. Aryl esters of phosphorus groups,especially phenyl esters, are reported to enhance oral bioavailability(DeLambert et al (1994) J. Med. Chem. 37: 498). Phenyl esters containinga carboxylic ester ortho to the phosphate have also been described(Khamnei and Torrence, (1996) J. Med. Chem. 39:4109-4115). Benzyl estersare reported to generate the parent phosphonic acid. In some cases,substituents at the ortho- or para-position may accelerate thehydrolysis. Benzyl analogs with an acylated phenol or an alkylatedphenol may generate the phenolic compound through the action of enzymes,e.g. esterases, oxidases, etc., which in turn undergoes cleavage at thebenzylic C—O bond to generate the phosphoric acid and the quinonemethide intermediate. Examples of this class of prodrugs are describedby Mitchell et al (1992) J. Chem. Soc. Perkin Trans. I2345; Brook et alWO 91/19721. Still other benzylic prodrugs have been describedcontaining a carboxylic ester-containing group attached to the benzylicmethylene (Glazier et al WO 91/19721). Thio-containing prodrugs arereported to be useful for the intracellular delivery of phosphonatedrugs. These proesters contain an ethylthio group in which the thiolgroup is either esterified with an acyl group or combined with anotherthiol group to form a disulfide. Deesterifecation or reduction of thedisulfide generates the free thio intermediate which subsequently breaksdown to the phosphoric acid and episulfide (Puech et al (1993) AntiviralRes., 22: 155-174; Benzaria et al (1996) J. Med. Chem. 39: 4958). Cyclicphosphonate esters have also been described as prodrugs ofphosphorus-containing compounds.

Prodrug esters in accordance with the invention are independentlyselected from the following groups: (1) mono-, di-, and tri-phosphateesters of tenofovir or emtricitabine or any other compound which uponadministration to a human subject is capable of providing (directly orindirectly) said mono-, di, or triphosphate ester; (2) carboxylic acidesters (3) sulphonate esters, such as alkyl- or aralkylsulphonyl (forexample, methanesulphonyl); (4) amino acid esters (for example, alanine,L-valyl or L-isoleucyl); (5) phosphonate; and (6) phosphonamidateesters.

Ester groups (1)-(6) may be substituted with; straight or branched chainC₁-C₁₈ alkyl (for example, methyl, n-propyl, t-butyl, or n-butyl);C₃-C₁₂ cycloalkyl; alkoxyalkyl (for example, methoxymethyl); arylalkyl(for example, benzyl); aryloxyalkyl (for example, phenoxymethyl); C₅-C₂₀aryl (for example, phenyl optionally substituted by, for example,halogen, C₁-C₄ alkyl, C₁-C₄ alkoxy, or amino; acyloxymethyl esters—CH₂C(═O)R⁹ (e.g. POM) or acyloxymethyl carbonates —CH₂C(═O)OR⁹ (e.g.POC) where R⁹ is C₁-C₆ alkyl, C₁-C₆ substituted alkyl, C₆-C₂₀ aryl orC₆-C₂₀ substituted aryl. For example, ester groups may be:—CH₂OC(O)C(CH₃)₃, —CH₂OC(O)OC(CH₃)₃ or —CH₂C(═O)OCH(CH₃)₂.

An exemplary aryl moiety present in such esters comprises a phenyl orsubstituted phenyl group. Many phosphate prodrug moieties are describedin U.S. Pat. No. 6,312,662; Jones et al (1995) Antiviral Research27:1-17; Kucera et al (1990) AIDS Res. Hum. Retro Viruses 6:491-501;Piantadosi et al (1991) J. Med. Chem. 34:1408-14; Hosteller et al (1992)Antimicrob. Agents Chemother. 36:2025-29; Hostetler et al (1990) J.Biol. Chem. 265:611127; and Siddiqui et al (1999) J. Med. Chem.42:4122-28.

Pharmaceutically acceptable prodrugs refer to a compound that ismetabolized in the host, for example hydrolyzed or oxidized, by eitherenzymatic action or by general acid or base solvolysis, to form anactive ingredient. Typical examples of prodrugs of the activeingredients of the combinations of the invention have biologicallylabile protecting groups on a functional moiety of the active compound.Prodrugs include compounds that can be oxidized, reduced, aminated,deaminated, esterified, deesterified, alkylated, dealkylated, acylated,deacylated, phosphorylated, dephosphorylated, or other functional groupchange or conversion involving forming or breaking chemical bonds on theprodrug.

Chemical Stability of a Pharmaceutical Formulation

The chemical stability of the active ingredients in a pharmaceuticalformulation is of concern to minimize the generation of impurities andensure adequate shelf-life. The active ingredients, tenofovir disoproxilfumarate and emtricitabine, in the pharmaceutical formulations of theinvention have relatively low pKa values, indicative of the potential tocause acidic hydrolysis of the active ingredients. Emtricitabine, with apKa of 2.65 (Emtriva™ Product Insert, Gilead Sciences, Inc. 2003,available at gilead.com) is subject to hydrolytic deamination of the5-fluoro cytosine nucleobase to form the 5-fluoro uridine nucleobase.Tenofovir disoproxil fumarate, with a pKa of 3.75 (Yuan L. et al“Degradation Kinetics of Oxycarbonyloxymethyl Prodrugs of Phosphonatesin Solution”, Pharmaceutical Research (2001) Vol. 18, No. 2, 234-237),is subject also to hydrolytic deamination of the exocyclic amine of theadenine nucleobase, and to hydrolysis of one or both of the POC estergroups (U.S. Pat. No. 5,922,695). It is desirable to formulate atherapeutic combination of tenofovir disoproxil fumarate andemtricitabine, and the physiological functional derivatives thereof,with a minimum of impurities and adequate stability.

The combinations of the present invention provide combinationpharmaceutical dosage forms which are chemically stable to aciddegradation of: (1) a first component (such as tenofovir disoproxilfumarate, and physiological functional derivatives; (2) a secondcomponent (such as emtricitabine, and physiological functionalderivatives; and (3) optionally a third component having antiviralactivity. The third component includes anti-HIV agents and include:protease inhibitors (PI), nucleoside reverse transcriptase inhibitors(NRTI), non-nucleoside reverse transcriptase inhibitors (NNRTI), andintegrase inhibitors. Exemplary third active ingredients to beadministered in combination with first and second components are shownin Table A. First and second components are as defined in the abovesection entitled: ACTIVE INGREDIENTS OF THE COMBINATIONS.

Salts

Any reference to any of the compounds in the compositions of theinvention also includes any physiologically acceptable salt thereof.Examples of physiologically acceptable salts of tenofovir DF,emtricitabine and their physiologically functional derivatives includesalts derived from an appropriate base, such as an alkali metal (forexample, sodium), an alkaline earth (for example, magnesium), ammoniumand NX₄ ⁺ (wherein X is C₁-C₄ alkyl), or an organic acid such as fumaricacid, acetic acid, succinic acid. Physiologically acceptable salts of anhydrogen atom or an amino group include salts of organic carboxylicacids such as acetic, benzoic, lactic, fumaric, tartaric, maleic,malonic, malic, isethionic, lactobionic and succinic acids; organicsulfonic acids, such as methanesulfonic, ethanesulfonic, benzenesulfonicand p-toluenesulfonic acids; and inorganic acids, such as hydrochloric,sulfuric, phosphoric and sulfamic acids. Physiologically acceptablesalts of a compound of an hydroxy group include the anion of saidcompound in combination with a suitable cation such as Na⁺ and NX₄ ⁺(wherein X is independently selected from H or a C₁-C₄ alkyl group).

For therapeutic use, salts of active ingredients of the combinations ofthe invention will be physiologically acceptable, i.e. they will besalts derived from a physiologically acceptable acid or base. However,salts of acids or bases which are not physiologically acceptable mayalso find use, for example, in the preparation or purification of aphysiologically acceptable compound. All salts, whether or not derivedfrom a physiologically acceptable acid or base, are within the scope ofthe present invention.

Administration of the Formulations

While it is possible for the active ingredients of the combination to beadministered alone and separately as monotherapies, it is preferable toadminister them as a pharmaceutical co-formulation. A two-part orthree-part combination may be administered simultaneously orsequentially. When administered sequentially, the combination may beadministered in one, two, or three administrations.

Preferably, two-part or three-part combinations are administered in asingle pharmaceutical dosage form. More preferably, a two-partcombination is administered as a single oral dosage form and athree-part combination is administered as two identical oral dosageforms. Examples include a single tablet of tenofovir disoproxil fumarateand emtricitabine, or two tablets of tenofovir disoproxil fumarate,emtricitabine, and efavirenz.

It will be appreciated that the compounds of the combination may beadministered: (1) simultaneously by combination of the compounds in aco-formulation or (2) by alternation, i.e. delivering the compoundsserially, sequentially, in parallel or simultaneously in separatepharmaceutical formulations. In alternation therapy, the delay inadministering the second, and optionally a third active ingredient,should not be such as to lose the benefit of a synergistic therapeuticeffect of the combination of the active ingredients. By either method ofadministration (1) or (2), ideally the combination should beadministered to achieve peak plasma concentrations of each of the activeingredients. A one pill once-per-day regimen by administration of acombination co-formulation may be feasible for some HIV-positivepatients. Effective peak plasma concentrations of the active ingredientsof the combination will be in the range of approximately 0.001 to 100μM. Optimal peak plasma concentrations may be achieved by a formulationand dosing regimen prescribed for a particular patient. It will also beunderstood that tenofovir DF and emtricitabine, or the physiologicallyfunctional derivatives of either thereof, whether presentedsimultaneously or sequentially, may be administered individually, inmultiples, or in any combination thereof. In general, during alternationtherapy (2), an effective dosage of each compound is administeredserially, where in co-formulation therapy (1), effective dosages of twoor more compounds are administered together.

Formulation of the Combinations

When the individual components of the combination are administeredseparately they are generally each presented as a pharmaceuticalformulation. The references hereinafter to formulations refer unlessotherwise stated to formulations containing either the combination or acomponent compound thereof. It will be understood that theadministration of the combination of the invention by means of a singlepatient pack, or patient packs of each formulation, within a packageinsert diverting the patient to the correct use of the invention is adesirable additional feature of this invention. The invention alsoincludes a double pack comprising in association for separateadministration, formulations of tenofovir disoproxil fumarate andemtricitabine, or a physiologically functional derivative of either orboth thereof.

The combination therapies of the invention include: (1) a combination oftenofovir DF and emtricitabine or (2) a combination containing aphysiologically functional derivative of either or both thereof.

The combination may be formulated in a unit dosage formulationcomprising a fixed amount of each active pharmaceutical ingredient for aperiodic, e.g. daily, dose or subdose of the active ingredients.

Pharmaceutical formulations according to the present invention comprisea combination according to the invention together with one or morepharmaceutically acceptable carriers or excipients and optionally othertherapeutic agents. Pharmaceutical formulations containing the activeingredient may be in any form suitable for the intended method ofadministration. When used for oral use for example, tablets, troches,lozenges, aqueous or oil suspensions, dispersible powders or granules,emulsions, hard or soft capsules, syrups or elixirs may be prepared(Remington's Pharmaceutical Sciences (Mack Publishing Co., Easton, Pa.).Compositions intended for oral use may be prepared according to anymethod known to the art for the manufacture of pharmaceuticalcompositions and such compositions may contain one or more agentsincluding antioxidants, sweetening agents, flavoring agents, coloringagents and preserving agents, in order to provide a palatablepreparation. Tablets containing the active ingredient in admixture withnon-toxic pharmaceutically acceptable excipient which are suitable formanufacture of tablets are acceptable. These excipients may be, forexample, inert diluents, such as calcium or sodium carbonate, lactose,lactose monohydrate, croscarmellose sodium, povidone, calcium or sodiumphosphate; granulating and disintegrating agents, such as maize starch,or alginic acid; binding agents, such as cellulose, microcrystallinecellulose, starch, gelatin or acacia; and lubricating agents, such asmagnesium stearate, stearic acid or talc. Tablets may be uncoated or maybe coated by known techniques including microencapsulation to delaydisintegration and absorption in the gastrointestinal tract and therebyprovide a sustained action over a longer period. For example, a timedelay material such as glyceryl monostearate or glyceryl distearatealone or with a wax may be employed.

Formulations for oral use may be also presented as hard gelatin capsuleswhere the active ingredient is mixed with an inert solid diluent, forexample pregelatinized starch, calcium phosphate or kaolin, or as softgelatin capsules wherein the active ingredient is mixed with water or anoil medium, such as peanut oil, liquid paraffin or olive oil.

Aqueous suspensions of the invention contain the active materials inadmixture with excipients suitable for the manufacture of aqueoussuspensions. Such excipients include a suspending agent, such as sodiumcarboxymethylcellulose, methylcellulose, hydroxypropyl methylcelluose,sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia,and dispersing or wetting agents such as a naturally occurringphosphatide (e.g., lecithin), a condensation product of an alkyleneoxide with a fatty acid (e.g., polyoxyethylene stearate), a condensationproduct of ethylene oxide with a long chain aliphatic alcohol (e.g.,heptadecaethyleneoxycetanol), a condensation product of ethylene oxidewith a partial ester derived from a fatty acid and a hexitol anhydride(e.g., polyoxyethylene sorbitan monooleate). The aqueous suspension mayalso contain one or more preservatives such as ethyl or n-propylp-hydroxybenzoate, one or more coloring agents, one or more flavoringagents and one or more sweetening agents, such as sucrose, sucralose orsaccharin.

Oil suspensions may be formulated by suspending the active ingredient ina vegetable oil, such as arachis oil, olive oil, sesame oil or coconutoil, or in a mineral oil such as liquid paraffin. The oral suspensionsmay contain a thickening agent, such as beeswax, hard paraffin or cetylalcohol. Sweetening agents, such as those set forth above, and flavoringagents may be added to provide a palatable oral preparation. Thesecompositions may be preserved by the addition of an antioxidant such asascorbic acid, BHT, etc.

Dispersible powders and granules of the invention suitable forpreparation of an aqueous suspension by the addition of water providethe active ingredient in admixture with a dispersing or wetting agent, asuspending agent, and one or more preservatives. Suitable dispersing orwetting agents and suspending agents are exemplified by those disclosedabove. Additional excipients, for example sweetening, flavoring andcoloring agents, may also be present.

The pharmaceutical compositions of the invention may also be in the formof oil-in-water emulsions or liposome formulations. The oily phase maybe a vegetable oil, such as olive oil or arachis oil, a mineral oil,such as liquid paraffin, or a mixture of these. Suitable emulsifyingagents include naturally-occurring gums, such as gum acacia and gumtragacanth, naturally occurring phosphatides, such as soybean lecithin,esters or partial esters derived from fatty acids and hexitolanhydrides, such as sorbitan monooleate, and condensation products ofthese partial esters with ethylene oxide, such as polyoxyethylenesorbitan monooleate. The emulsion may also contain sweetening andflavoring agents. Syrups and elixirs may be formulated with sweeteningagents, such as glycerol, sorbitol or sucrose. Such formulations mayalso contain a demulcent, a preservative, a flavoring or a coloringagent.

The pharmaceutical compositions of the invention may be in the form of asterile injectable preparation, such as a sterile injectable aqueous oroleaginous suspension. This suspension may be formulated according tothe known art using those suitable dispersing or wetting agents andsuspending agents which have been mentioned above. The sterileinjectable preparation may also be a sterile injectable solution orsuspension in a non-toxic parenterally acceptable diluent or solvent,such as a solution in 1,3-butane-diol or prepared as a lyophilizedpowder. Among the acceptable vehicles and solvents that may be employedare water, Ringer's solution and isotonic sodium chloride solution. Inaddition, sterile fixed oils may conventionally be employed as a solventor suspending medium. For this purpose any bland fixed oil may beemployed including synthetic mono- or diglycerides. In addition, fattyacids such as oleic acid may likewise be used in the preparation ofinjectables.

The pharmaceutical compositions of the invention may be injectedparenterally, for example, intravenously, intraperitoneally,intrathecally, intraventricularly, intrasternally, intracranially,intramuscularly or subcutaneously, or they may be administered byinfusion techniques. They are best used in the form of a sterile aqueoussolution which may contain other substances, for example, enough saltsor glucose to make the solution isotonic with blood. The aqueoussolutions should be suitably buffered (preferably to a pH of from 3 to9), if necessary. The preparation of suitable parenteral formulationsunder sterile conditions is readily accomplished by standardpharmaceutical techniques well known to those skilled in the art.

The pharmaceutical compositions of the invention may also beadministered intranasally or by inhalation and are convenientlydelivered in the form of a dry powder inhaler or an aerosol spraypresentation from a pressurized container or a nebuliser with the use ofa suitable propellant, e.g. dichlorodifluoromethane,trichlorofluoromethane, dichlorotetrafluoroethane, a hydrofluoroalkanesuch as 1,1,1,2-tetrafluoroethane (HFC 134a), carbon dioxide or othersuitable gas. In the case of a pressurized aerosol, the dosage unit maybe determined by providing a valve to deliver a metered amount. Thepressurized container or nebuliser may contain a solution or suspensionof the composition, e.g. using a mixture of ethanol and the propellantas the solvent, which may additional contain a lubricant, e.g. sorbitantrioleate. Capsules and cartridges (made, for example, from gelatin) foruse in an inhaler or insufflator may be formulated to contain a powdermix of a compound of the formula (I) and a suitable powder base such aslactose or starch. Aerosol or dry powder formulations are preferablyarranged so that each metered dose or “puff” contains from 20 μg to 20mg of a composition for delivery to the patient. The overall daily dosewith an aerosol will be in the range of from 20 μg to 20 mg which may beadministered in a single dose or, more usually, in divided dosesthroughout the day.

The amount of active ingredient that may be combined with the carriermaterial to produce a single dosage form will vary depending upon thehost treated and the particular mode of administration. For example, atime-release formulation intended for oral administration to humans maycontain approximately 1 to 1000 mg of active material compounded with anappropriate and convenient amount of carrier material which may varyfrom about 5 to about 95% of the total compositions (weight:weight). Thepharmaceutical composition can be prepared to provide easily measurableamounts for administration. For example, an aqueous solution intendedfor intravenous infusion may contain from about 3 to 500 μg of theactive ingredient per milliliter of solution in order that infusion of asuitable volume at a rate of about 30 mL/hr can occur. As noted above,formulations of the present invention suitable for oral administrationmay be presented as discrete units such as capsules, cachets or tabletseach containing a predetermined amount of the active ingredient; as apowder or granules; as a solution or a suspension in an aqueous ornon-aqueous liquid; or as an oil-in-water liquid emulsion or awater-in-oil liquid emulsion. The active ingredient may also beadministered as a bolus, electuary or paste.

The combinations of the invention may conveniently be presented as apharmaceutical formulation in a unitary dosage form. A convenientunitary dosage formulation contains the active ingredients in any amountfrom 1 mg to 1 g each, for example but not limited to, 10 mg to 300 mg.The synergistic effects of tenofovir DF in combination withemtricitabine may be realized over a wide ratio, for example 1:50 to50:1 (tenofovir DF:emtricitabine). In one embodiment the ratio may rangefrom about 1:10 to 10:1. In another embodiment, the weight/weight ratioof tenofovir to emtricitabine in a co-formulated combination dosageform, such as a pill, tablet, caplet or capsule will be about 1, i.e. anapproximately equal amount of tenofovir DF and emtricitabine. In otherexemplary co-formulations, there may be more or less tenofovir than FTC.For example, 300 mg tenofovir DF and 200 mg emtricitabine can beco-formulated in a ratio of 1.5:1 (tenofovir DF: emtricitabine). In oneembodiment, each compound will be employed in the combination in anamount at which it exhibits antiviral activity when used alone.Exemplary Formulations A, B, C, D, E, and F (Examples) have ratios of12:1 to 1:1 (tenofovir F: emtricitabine). Exemplary Formulations A, B,C, D, E, and F use amounts of tenofovir DF and emtricitabine rangingfrom 25 mg to 300 mg. Other ratios and amounts of the compounds of saidcombinations are contemplated within the scope of the invention.

A unitary dosage form may further comprise tenofovir DF andemtricitabine, or physiologically functional derivatives of eitherthereof, and a pharmaceutically acceptable carrier.

It will be appreciated by those skilled in the art that the amount ofactive ingredients in the combinations of the invention required for usein treatment will vary according to a variety of factors, including thenature of the condition being treated and the age and condition of thepatient, and will ultimately be at the discretion of the attendingphysician or health care practitioner. The factors to be consideredinclude the route of administration and nature of the formulation, theanimal's body weight, age and general condition and the nature andseverity of the disease to be treated. For example, in a Phase I/IImonotherapy study of emtricitabine, patients received doses ranging from25 mg to 200 mg twice-a-day for two weeks. At each dose regimen greateror equal to 200 mg, a 98-percent (1.75 log 10) or greater viralsuppression was observed. A once-a-day dose of 200 mg of emtricitabinereduced the viral load by an average of 99 percent (1.92 log 10).Viread® (tenofovir DF) has been approved by the FDA for the treatmentand prophylaxis of HIV infection as a 300 mg oral tablet. Emtriva™(emtricitabine) has been approved by the FDA for the treatment of HIV asa 200 mg oral tablet.

It is also possible to combine any two of the active ingredients in aunitary dosage form for simultaneous or sequential administration with athird active ingredient. The three-part combination may be administeredsimultaneously or sequentially. When administered sequentially, thecombination may be administered in two or three administrations. Thirdactive ingredients have anti-HIV activity and include proteaseinhibitors (PI), nucleoside reverse transcriptase inhibitors (NRTI),non-nucleoside reverse transcriptase inhibitors (NNRTI), and integraseinhibitors. Exemplary third active ingredients to be administered incombination with tenofovir DF, emtricitabine, and their physiologicalfunctional derivatives, are shown in Table A.

TABLE A 5,6 dihydro-5-azacytidine 5-aza 2′deoxycytidine 5-azacytidine5-yl-carbocyclic 2′-deoxyguanosine (BMS200,475) 9(arabinofuranosyl)guanine; 9-(2′ deoxyribofuranosyl)guanine 9-(2′-deoxy2′fluororibofuranosyl)-2,6-diaminopurine 9-(2′-deoxy2′fluororibofuranosyl)guanine 9-(2′-deoxyribofuranosyl)-2,6diaminopurine 9-(arabinofuranosyl)-2,6 diaminopurine Abacavir, Ziagen ®Acyclovir, ACV; 9-(2-hydroxyethoxylmethyl)guanine Adefovir dipivoxil,Hepsera ® amdoxivir, DAPD Amprenavir, Agenerase ® araA;9-β-D-arabinofuranosyladenine (Vidarabine) atazanivir sulfate(Reyataz ®) AZT; 3′-azido-2′,3′-dideoxythymdine, Zidovudine,(Retrovir ®) BHCG;(.+−.)-(1a,2b,3a)-9-[2,3-bis(hydroxymethyl)cyclobutyl]guanineBMS200,475; 5-yl-carbocyclic 2′-deoxyguanosine Buciclovir; (R)9-(3,4-dihydroxybutyl)guanine BvaraU;1-β-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil (Sorivudine) CalanolideA Capravirine CDG; carbocyclic 2′-deoxyguanosine Cidofovir, HPMPC;(S)-9-(3-hydroxy-2- phosphonylmethoxypropyl)cytosine Clevudine, L-FMAU;2′-Fluoro-5-methyl-β-L-arabino-furanosyluracil Combivir ®(lamivudine/zidovudine) Cytallene;[1-(4′-hydroxy-1′,2′-butadienyl)cytosine] d4C;3′-deoxy-2′,3′-didehydrocytidine DAPD; (−)-β-D-2,6-diaminopurinedioxolane ddA; 2′,3′-dideoxyadenosine ddAPR;2,6-diaminopurine-2′,3′-dideoxyriboside ddC; 2′,3′-dideoxycytidine(Zalcitabine) ddI; 2′,3′-dideoxyinosine, didanosine, (Videx ®, Videx ®EC) Delavirdine, Rescriptor ® Didanosine, ddI, Videx ®;2′,3′-dideoxyinosine DXG; dioxolane guanosineE-5-(2-bromovinyl)-2′-deoxyuridine Efavirenz, Sustiva ® Enfuvirtide,Fuzeon ® F-ara-A; fluoroarabinosyladenosine (Fludarabine) FDOC;(−)-β-D-5-fluoro-1-[2-(hydroxymethyl)-1,3-dioxolane]cytosine FEAU;2′-deoxy-2′-fluoro-1-β-D-arabinofuranosyl-5-ethyluracil FIAC;1-(2-deoxy-2-fluoro-β-D-arabinofuranosyl)-5-iodocytosine FIAU;1-(2-deoxy-2-fluoro-β-D-arabinofuranosyl)-5-iodouridine FLG;2′,3′-dideoxy-3′-fluoroguanosine FLT; 3′-deoxy-3′-fluorothymidineFludarabine; F-ara-A; fluoroarabinosyladenosine FMAU;2′-Fluoro-5-methyl-β-L-arabino-furanosyluracil FMdC Foscarnet;phosphonoformic acid, PFA FPMPA;9-(3-fluoro-2-phosphonylmethoxypropyl)adenine Gancyclovir, GCV;9-(1,3-dihydroxy-2-propoxymethyl)guanine GS-7340;9-[R-2-[[(S)-[[(S)-1-(isopropoxycarbonyl)ethyl]amino]-  phenoxyphosphinyl]methoxy]propyl]adenine HPMPA;(S)-9-(3-hydroxy-2-phosphonylmethoxypropyl)adenine HPMPC;(S)-9-(3-hydroxy-2-phosphonylmethoxypropyl)cytosine (Cidofovir)Hydroxyurea, Droxia ® Indinavir, Crixivan ® Kaletra ®(lopinavir/ritonavir) Lamivudine, 3TC, Epivir ™;(2R,5S,cis)-4-amino-1-(2-  hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-pyrimidin-2-one L-d4C;L-3′-deoxy-2′,3′-didehydrocytidine L-ddC; L-2′,3′-dideoxycytidineL-Fd4C; L-3′-deoxy-2′,3′-didehydro-5-fluorocytidine L-FddC;L-2′,3′-dideoxy-5-fluorocytidine Lopinavir Nelfinavir, Viracept ®Nevirapine, Viramune ® Oxetanocin A;9-(2-deoxy-2-hydroxymethyl-β-D-erythro- oxetanosyl)adenine Oxetanocin G;9-(2-deoxy-2-hydroxymethyl-β-D-erythro- oxetanosyl)guanine PenciclovirPMEDAP; 9-(2-phosphonylmethoxyethyl)-2,6-diaminopurine PMPA, tenofovir;(R)-9-(2-phosphonylmethoxypropyl)adenine PPA; phosphonoacetic acidRibavirin; 1-β-D-ribofuranosyl-1,2,4-triazole-3-carboxamide Ritonavir,Norvir ® Saquinavir, Invirase ®, Fortovase ® Sorivudine, BvaraU;1-β-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil Stavudine, d4T, Zerit ®;2′,3′-didehydro-3′-deoxythymidine Trifluorothymidine, TFT;Trifluorothymidine Trizivir ® (abacavir sulfate/lamivudine/zidovudine)Vidarabine, araA; 9-β-D-arabinofuranosyladenine Zalcitabine, Hivid ®,ddC; 2′,3′-dideoxycytidine Zidovudine, AZT, Retrovir ®;3′-azido-2′,3′-dideoxythymdine Zonavir; 5-propynyl-1-arabinosyluracil

Another aspect of the present invention is a three-part combinationcomprising tenofovir DF, FTC, and9-[(R)-2-[[(S)-[[(S)-1-(isopropoxycarbonyl)ethyl]amino]phenoxyphosphinyl]methoxy]propyl]adenine,also designated herein as GS-7340, which has the structure:

GS-7340 is a prodrug of tenofovir and the subject of commonly owned,pending application, U.S. Ser. No. 09/909,560, filed Jul. 20, 2001 andBecker et al WO 02/08241.

For example, a ternary unitary dosage may contain 1 mg to 1000 mg oftenofovir disoproxil fumarate, 1 mg to 1000 mg of emtricitabine, and 1mg to 1000 mg of the third active ingredient. As a further feature ofthe present invention, a unitary dosage form may further comprisetenofovir DF, emtricitabine, the third active ingredient, orphysiologically functional derivatives of the three active ingredientsthereof, and a pharmaceutically acceptable carrier.

Combinations of the present invention enable patients greater freedomfrom multiple dosage medication regimens and ease the needed diligencerequired in remembering and complying with complex daily dosing timesand schedules. By combining tenofovir disoproxil fumarate andemtricitabine into a single dosage form, the desired daily regimen maybe presented in a single dose or as two or more sub-doses per day. Thecombination of co-formulated tenofovir DF and emtricitabine may beadministered as a single pill, once per day.

A further aspect of the invention is a patient pack comprising at leastone active ingredient: tenofovir disoproxil fumarate, emtricitabine, ora physiologically functional derivative of either of the combination andan information package or product insert containing directions on theuse of the combination of the invention.

Segregation of active ingredients in pharmaceutical powders andgranulations is a widely recognized problem that can result ininconsistent dispersions of the active ingredients in final dosageforms. Some of the main factors contributing to segregation are particlesize, shape and density. Segregation is particularly troublesome whenattempting to formulate a single homogenous tablet containing multipleactive ingredients having different densities and different particlesizes. Glidants are substances that have traditionally been used toimprove the flow characteristics of granulations and powders by reducinginterparticulate friction. See Lieberman, Lachman, & Schwartz,Pharmaceutical Dosage Forms: Tablets, Volume 1, p. 177-178 (1989),incorporated herein by reference. Glidants are typically added topharmaceutical compositions immediately prior to tablet compression tofacilitate the flow of granular material into the die cavities of tabletpresses. Glidants include: colloidal silicon dioxide, asbestos freetalc, sodium aluminosilicate, calcium silicate, powdered cellulose,microcrystalline cellulose, corn starch, sodium benzoate, calciumcarbonate, magnesium carbonate, metallic stearates, calcium stearate,magnesium stearate, zinc stearate, stearowet C, starch, starch 1500,magnesium lauryl sulfate, and magnesium oxide. Exemplary TabletFormulation A has colloidal silicon dioxide (Examples). Glidants can beused to increase and aid blend composition homogeneity in formulationsof anti-HIV drugs (U.S. Pat. No. 6,113,920). The novel compositions ofthe present invention may contain glidants to effect and maintainhomogeneity of active ingredients during handling prior to tabletcompression.

The present invention provides pharmaceutical formulations combining theactive ingredients tenofovir DF and emtricitabine, or physiologicallyfunctional derivatives thereof in a sufficiently homogenized form, and amethod for using this pharmaceutical formulation. An object of thepresent invention is to utilize glidants to reduce the segregation ofactive ingredients in pharmaceutical compositions during pre-compressionmaterial handling. Another object of the present invention is to providea pharmaceutical formulation combining the active ingredients tenofovirDF and emtricitabine, or physiologically functional derivatives thereof,with a pharmaceutically acceptable glidant, resulting in a mixturecharacterized by a pharmaceutically acceptable measure of homogeneity.

Formulations include those suitable for oral, rectal, nasal, topical(including transdermal, buccal and sublingual), vaginal or parenteral(including subcutaneous, intramuscular, intravenous and intradermal)administration. The formulations may conveniently be presented in unitdosage form and may be prepared by any methods well known in the art ofpharmacy. Such methods represent a further feature of the presentinvention and include the step of bringing into association the activeingredients with the carrier, which constitutes one or more accessoryingredients, and maintaining chemical stability. In general, theformulations are prepared by uniformly and intimately bringing intoassociation the active ingredients with liquid carriers or finelydivided solid carriers or both, and then if necessary shaping theproduct.

Formulations of the present invention suitable for oral administrationmay be presented as discrete units such as capsules, caplets, cachets ortablets each containing a predetermined amount of the activeingredients; as a powder or granules; as a solution or a suspension inan aqueous or non-aqueous liquid; or as an oil-in-water liquid emulsionor a water-in-oil liquid emulsion. The active ingredient may also bepresented as a bolus, electuary or paste.

A tablet may be made by compression or molding, optionally with one ormore accessory ingredients. Compressed tablets may be prepared bycompressing in a suitable machine the active ingredients in afree-flowing form such as a powder or granules, optionally mixed with abinder (e.g. povidone, gelatin, hydroxypropyl methylcellulose),lubricant, inert diluent, preservative, disintegrant (e.g. sodium starchglycollate, cross-linked povidone, cross-linked sodium carboxymethylcellulose) surface-active or dispersing agent. Molded tablets may bemade by molding a mixture of the powdered compound moistened with aninert liquid diluent in a suitable machine. The tablets may optionallybe coated or scored and may be formulated so as to provide slow orcontrolled release of the active ingredients therein using, for example,cellulose ether derivatives (e.g., hydroxypropyl methylcellulose) ormethacrylate derivatives in varying proportions to provide the desiredrelease profile. Tablets may optionally be provided with an entericcoating, to provide release in parts of the gut other than the stomach.

Formulations suitable for topical administration in the mouth includelozenges comprising the active ingredients in a flavored base, usuallysucrose and acacia or tragacanth; pastiles comprising the activeingredient in an inert basis such as gelatin and glycerin, or sucroseand acacia; and mouthwashes comprising the active ingredient in asuitable liquid carrier. Formulations for rectal administration may bepresented as a suppository with a suitable base comprising, for example,cocoa butter or a salicylates. Topical administration may also be bymeans of a transdermal iontophoretic device.

Formulations suitable for vaginal administration may be presented aspessaries, tampons, creams, gels, pastes, foams or spray formulationscontaining in addition to the active ingredient such carriers as areknown in the art to be appropriate.

Formulations suitable for penile administration for prophylactic ortherapeutic use may be presented in condoms, creams, gels, pastes, foamsor spray formulations containing in addition to the active ingredientsuch carriers as are known in the art to be appropriate.

Pharmaceutical formulations suitable for rectal administration whereinthe carrier is a solid are most preferably presented as unit dosesuppositories. Suitable carriers include cocoa butter and othermaterials commonly used in the art. The suppositories may beconveniently formed by admixture of the active combination with thesoftened or melted carrier(s) followed by chilling and shaping inmoulds.

Formulations suitable for parenteral administration include aqueous andnonaqueous isotonic sterile injection solutions which may containanti-oxidants, buffers, bacteriostats and solutes which render theformulation isotonic with the blood of the intended recipient; andaqueous and non-aqueous sterile suspensions which may include suspendingagents and thickening agents; and liposomes or other microparticulatesystems which are designed to target the compound to blood components orone or more organs. The formulations may be presented in unit-dose ormulti-dose sealed containers, for example, ampoules and vials, and maybe stored in a freeze-dried (lyophilized) condition requiring only theaddition of the sterile liquid carrier, for example water for injection,immediately prior to use. Extemporaneous injection solutions andsuspensions may be prepared from sterile powders, granules and tabletsof the kind previously described.

Exemplary unit dosage formulations are those containing a daily dose ordaily subdose of the active ingredients, as hereinbefore recited, or anappropriate fraction thereof. It should be understood that in additionto the ingredients particularly mentioned above the formulations of thisinvention may include other agents conventional in the art having regardto the type of formulation in question, for example, those suitable fororal administration may include such further agents as sweeteners,thickeners and flavoring agents.

The compounds of the combination of the present invention may beobtained in a conventional manner, known to those skilled in the art.Tenofovir disoproxil fumarate can be prepared, for example, as describedin U.S. Pat. No. 5,977,089. Methods for the preparation of FTC aredescribed in WO 92/14743, incorporated herein by reference.

Composition Use

Compositions of the present invention are administered to a human orother mammal in a safe and effective amount as described herein. Thesesafe and effective amounts will vary according to the type and size ofmammal being treated and the desired results of the treatment. Any ofthe various methods known by persons skilled in the art for packagingtablets, caplets, or other solid dosage forms suitable for oraladministration, that will not degrade the components of the presentinvention, are suitable for use in packaging. The combinations may bepackaged in glass and plastic bottles. Tablets, caplets, or other soliddosage forms suitable for oral administration may be packaged andcontained in various packaging materials optionally including adessicant e.g. silica gel. Packaging may be in the form of unit doseblister packaging. For example, a package may contain one blister trayof tenofovir DF and another blister tray of emtricitabine pills,tablets, caplets, or capsule. A patient would take one dose, e.g. apill, from one tray and one from the other. Alternatively, the packagemay contain a blister tray of the co-formulated combination of tenofovirDF and emtricitabine in a single pill, tablet, caplet or capsule. As inother combinations and packaging thereof, the combinations of theinvention include physiological functional derivatives of tenofovir DFand FTC.

The packaging material may also have labeling and information related tothe pharmaceutical composition printed thereon. Additionally, an articleof manufacture may contain a brochure, report, notice, pamphlet, orleaflet containing product information. This form of pharmaceuticalinformation is referred to in the pharmaceutical industry as a “packageinsert.” A package insert may be attached to or included with apharmaceutical article of manufacture. The package insert and anyarticle of manufacture labeling provides information relating to thepharmaceutical composition. The information and labeling providesvarious forms of information utilized by health-care professionals andpatients, describing the composition, its dosage and various otherparameters required by regulatory agencies such as the United StatesFood and Drug Agency.

Assays of the Combinations

The combinations of the inventions may be tested for in vitro activityagainst HIV and sensitivity, and for cytotoxicity in laboratory adaptedcell lines, e.g. MT2 and in peripheral blood mononuclear cells (PBMC)according to standard assays developed for testing anti-HIV compounds,such as WO 02/068058 and U.S. Pat. No. 6,475,491. Combination assays maybe performed at varying concentrations of the compounds of thecombinations to determine EC₅₀ by serial dilutions.

EXEMPLARY FORMULATIONS

The following examples further describe and demonstrate particularembodiments within the scope of the present invention. Techniques andformulations generally are found in Remington's Pharmaceutical Sciences(Mack Publishing Co., Easton, Pa.). The examples are given solely forillustration and are not to be construed as limitations as manyvariations are possible without departing from spirit and scope of theInvention. The following examples are intended for illustration only andare not intended to limit the scope of the invention in any way. “Activeingredient” denotes tenofovir disoproxil fumarate, emtricitabine, or aphysiologically functional derivative of either thereof.

Tablet Formulation

The following exemplary formulations A, B, C, D, E, and F are preparedby wet granulation of the ingredients with an aqueous solution, additionof extragranular components and then followed by addition of magnesiumstearate and compression.

mg/tablet Formulation A: Tenofovir Disoproxil Fumarate 300 emtricitabine200 Microcrystalline Cellulose 200 Lactose Monohydrate 175Croscarmellose Sodium 60 Pregelatinized Starch 50 Colloidal silicondioxide 5 Magnesium Stearate 10 total: 1000 Formulation B: TenofovirDisoproxil fumarate 300 emtricitabine 100 Microcrystalline Cellulose 200Lactose Monohydrate 180 Sodium Starch Glycollate 60 PregelatinizedStarch 50 Magnesium Stearate 10 total: 900 Formulation C: TenofovirDisoproxil fumarate 200 emtricitabine 200 Microcrystalline Cellulose 200Lactose Monohydrate 180 Sodium Starch Glycollate 60 PregelatinizedStarch 50 Magnesium Stearate 10 total: 900 Formulation D: TenofovirDisoproxil fumarate 300 emtricitabine 25 Microcrystalline Cellulose 200Lactose Monohydrate 180 Sodium Starch Glycollate 60 PregelatinizedStarch 50 Magnesium Stearate 10 total: 825 Formulation E: TenofovirDisoproxil fumarate 200 emtricitabine 25 Microcrystalline Cellulose 200Lactose Monohydrate 180 Sodium Starch Glycollate 60 PregelatinizedStarch 50 Magnesium Stearate 10 total: 725 Formulation F: TenofovirDisoproxil fumarate 100 emtricitabine 100 Microcrystalline Cellulose 200Lactose Monohydrate 180 Sodium Starch Glycollate 60 PregelatinizedStarch 50 Magnesium Stearate 10 total: 700Formulation G (Controlled Release Formulation):This formulation is prepared by wet granulation of the ingredients withan aqueous solution, followed by the addition of magnesium stearate andcompression.

mg/tablet Tenofovir Disoproxil fumarate 300 emtricitabine 200Hydroxypropyl Methylcellulose 112 Lactose B.P. 53 Pregelatinized StarchB.P. 28 Magnesium Stearate total: 700Drug release takes place over a period of about 6-8 hours and iscomplete after 12 hours.Capsule FomulationsFormulation H:A capsule formulation is prepared by admixing the ingredients andfilling into a two-part hard gelatin or hydroxypropyl methylcellulosecapsule.

mg/capsule Active Ingredient 500 Microcrystalline Cellulose 143 SodiumStarch Glycollate 25 Magnesium Stearate 2 total: 670Formulation I (Controlled Release Capsule):The following controlled release capsule formulation is prepared byextruding ingredients a, b, and c using an extruder, followed byspheronization of the extrudate and drying. The dried pellets are thencoated with release-controlling membrane (d) and filled into atwo-piece, hard gelatin or hydroxypropyl methylcellulose capsule.

mg/capsule (a) Active Ingredient 500 (b) Microcrystalline Cellulose 125(c) Lactose B.P. 125 (d) Ethyl Cellulose 13 total: 763Formulation J (Oral Suspension):The active ingredients are admixed with the ingredients and filling themas dry powder.Purified water is added and mixed well before use.

Active Ingredient 500 mg Confectioner's Sugar 2000 mg Simethicone 300 mgMethylparaben 30 mg Propylparaben 10 mg Flavor, Peach 500 mg PurifiedWater q.s. to 5.00 mlFormulation K (Suppository):One-fifth of the Witepsol H15 is melted in a steam-jacketed pan at 45°C. maximum. The active ingredients are sifted through a 200 micron sieveand added to the molten base with mixing, using a Silverson fitted witha cutting head, until a smooth dispersion is achieved. Maintaining themixture at 45° C., the remaining Witepsol H15 is added to the suspensionand stirred to ensure a homogenous mix. The entire suspension is passedthrough a 250 micron stainless steel screen and, with continuousstirring, is allowed to cool to 40° C. At a temperature of 38° C. to 40°C., 2.02 g of the mixture is filled into suitable, 2 ml plastic molds.The suppositories are allowed to cool to room temperature.

mg/Suppository Active Ingredient 500 Hard Fat, B.P. (WitepsolH15-Dynamit Nobel) 1770 total 2270Fixed Dose Combination Tablet

A fixed dose combination tablet of tenofovir disoproxil fumarate (TDF)300 mg/emtricitabine 200 mg was formulated using a wetgranulation/fluid-bed drying process using conventional methods. See:U.S. Pat. No. 5,935,946; L. Young (editor). Tableting SpecificationManual 5^(th) ed., American Pharmaceutical Association, Washington,D.C., (2001); L. Lachman, H. Lieberman (editors). Pharmaceutical DosageForms: Tablets (Vol 2), Marcel Dekker Inc., New York, 185-202 (1981); J.T. Fell and J. M. Newton, J. Pharm. Pharmacol. 20, 657-659 (1968); USPharmacopeia 24-National Formulary 19, “Tablet Friability”, Chapter<1216>, Page 2148 (2000).

The effects of granulation water level (ranging from 40% to 50% w/w) andwet massing time were studied on the physicochemical properties of thefinal powder blend and its performance with respect to blend uniformityand compressibility (tablet compactibility). In addition, contentuniformity, assay, stability and dissolution performance was evaluatedfor the TDF/emtricitabine fixed dose combination tablets.

Formulation Equipment

Equipment included a high shear mixer equipped with a pressure tank andspray nozzle tip to add the granulating water, a fluid-bed dryer, amill, a tumble blender, a rotary tablet press, and a tablet deduster.

Formulation Process

The dried, milled powder was blended with the extragranularmicrocrystalline cellulose and croscarmellose sodium and then blendedwith magnesium stearate. Powder samples were removed after mixing withthe magnesium stearate. The blend samples were evaluated for, bulkdensity, mesh analysis and compressibility. The powder blend mixed withthe magnesium stearate was compressed into tablets on a press setup.

Materials

The following Table 1 lists the quantitative composition of theTDF/emtricitabine tablet formulation.

TABLE 1 Unit Formula for Quantity per tablet cores 12 kg BatchIngredient % w/w (mg/tablet) (kg) Tenofovir Disoproxil 30.0 300.0 3.60Fumarate^(a) Emtricitabine^(a) 20.0 200.0 2.40 Pregelatinized Starch,NF/EP 5.0 50.0 0.60 Croscarmellose Sodium, 6.0 60.0 0.72 NF/EP LactoseMonohydrate, NF/EP^(a) 8.0 80.0 0.96 Microcrystalline Cellulose, 30.0300.0 3.60 NF/EP^(c) Magnesium Stearate, NF/EP 1.0 10.0 0.12 PurifiedWater, USP/EP ^(b) ^(b) ^(b) Totals 100.0 1000.0 12.00 ^(a)Actual weightis adjusted based on the Drug Content Factor (DCF) of tenofovirdisoproxil fumarate and emtricitabine. ^(b)Water removed during drying.Characterization Equipment

Moisture content was measured by loss on drying using a heatlamp/balance system. The powder blend was sampled with a sampling thieffitted with chambers to determine powder blend uniformity. Duplicatesamples were removed from each of several locations in the blender.Blend uniformity analysis was performed on one sample from eachlocation.

Particle size analysis of the final powder blend was determined bysifting a multi-gram sample through a screen using a sonic sifter. Thequantity of final powder blend retained on each sieve and the finescollector was determined by calculating the difference in weight betweenthe sieves and fines collector before and after the test. The geometricmean diameter particle size was calculated by logarithmic weighting ofthe sieved distribution.

Bulk density was determined by filling a graduated cylinder with thefinal powder blend and measuring the weight differential between theempty and filled graduate cylinder per unit volume.

Tablets were characterized for friability using a friabilator, ahardness tester, a thickness micrometer equipped with a printer, and aweighing balance.

Compression characteristics were determined using a rotary tablet pressequipped with a flat-faced, beveled edged punch to a target weight of400 mg. The powder blends were compressed using target upper punchpressures ranging from approximately 100 to 250 MPa. The apparentnormalized ejection force was determined and normalized for tabletthickness and diameter.

Tablet hardness was determined using a hardness tester. Tablet thicknesswas determined using a micrometer, and tablet weights were determinedusing a top loading balance.

Wet Granulation

The powders were blended in a granulator and then granulated usingwater. The impeller and chopper speeds were kept constant in the blenderat a low setting during the granulation and wet massing operations.After water addition, the impeller and chopper were stopped and thegranulator bowl was opened to observe the granulation consistency andtexture. The lid was closed and the wet massing phase was performed.Acceptable granules had 40% w/w and 60% w/w water, respectively.

Wet Milling

To facilitate a uniform drying process, each wet granulation wasdeagglomerated with a mill fitted with a screen and an impeller. Themilled wet granules were charged into a fluid-bed dryer immediatelyfollowing wet milling.

Fluid-Bed Drying

Milled wet granules were dried using an inlet air setpoint temperatureof about 70° C. and airflow of approximately 100 cfm. The target LOD wasabout 1.0% with a range of not more than (NMT) 1.5%. The total fluid-beddrying time ranged from 53 to 75 minutes. Final LOD ranged from 0.4% to0.7% for all of the batches dried. The final exhaust temperatures forall the batches ranged from 47° C. to 50° C.

Dry Milling

All dried granules were milled through a perforated screen. The mill wasequipped with a square impeller and operated. The lots were milled andmanually transferred to the V-blender.

Blending

Each lot was blended using the V-blender. In one set of threeformulations, starting with 12 kg materials, final powder blend yieldavailable for compression after blending ranged from 10.5 kg (87.5%) to11.1 kg (92.5%). The final powder blend bulk density ranged from 0.48 to0.58 g/cc and the geometric mean diameter particle size ranged from 112to 221 μm. Percent water and wet massing time affect final powder blendparticle size and bulk density.

The powder blending for both tenofovir DF and emtricitabine gave a mean(n=10) strength value for tenofovir DF ranged from 100.6% to 102.8% oftarget strength for the lots and the relative standard deviation (RSD)was from 0.5% to 1.7%. The mean (n=10) strength value for emtricitabineranged from 101.3% to 104.1% of target strength for the lots with therelative standard deviation (RSD) ranged from 0.6% to 1.7%. The finalpowder blend moisture level ranged from 0.8% to 1.1% LOD.

Tablet Compression

The final blends were compressed using a rotary tablet press and thetablets were film-coated.

Three 300 gm formulations (Table 2) were granulated in a granulatorequipped with a 1-L bowl. The quantities of intragranular componentswere based on a 300 g total batch size. The formulations in lots 1 and 2differed in the amount of microcrystalline cellulose 30% vs. 20% w/w,respectively. Lots 2 and 3 were identical except for the type of binder.Lot 2 contained 5% w/w of pregelatinized starch and lot 3 contained 5%w/w povidone as binder.

TABLE 2 Ingredient Lot 1 % w/w Lot 2 % w/w Lot 3 % w/w TenofovirDisoproxil 30.0 30.0 30.0 Fumarate Emtricitabine 20.0 20.0 20.0Pregelatinized Starch, 5.0 5.0 N/A NF/EP Povidone, USP/NF (C- N/A N/A5.0 30) Croscarmellose 6.0 6.0 6.0 Sodium, NF/EP Lactose Monohydrate,8.0 18.0 18.0 NF/EP Microcrystalline 30.0 20.0 20.0 Cellulose, NF/EP^(a)Magnesium Stearate, 1.0 1.0 1.0 NF/EP Purified Water, ^(a) ^(a) ^(a)USP/EP Total 100.0 100.0 100.0 ^(a)Water removed during drying.

After water addition, the impeller and chopper were stopped and thegranulator bowl was opened to observe the granulation consistency andtexture. To achieve similar granulation consistency, lots 1, 2, and 3were granulated with 45%, 40%, and 30% w/w water, respectively. The lidwas closed and the wet massing phase was performed. All lots had a 30sec wet massing resulting in acceptable granulations. The wetgranulations from all batches were hand screened through a sieve todeagglomerate. The resulting granulations were tray dried in aconvection oven set at 60° C. for approximately 20 hours to an LOD<1.0%. The dried granulations from all batches were hand screenedthrough a sieve. In order to fit the granulation into the small scale(300 mL) V-blender, the final blend batch size was adjusted to 100 g. Aportion, 81 g of the resulting blend from Lot 1 was blended with 15 gmicrocrystalline cellulose, 3 g croscarmellose sodium and 1 g magnesiumstearate. 86 g of the resulting granulation from Lot 2 and Lot 3 wereeach blended with 10 g microcrystalline cellulose, 3 g croscarmellosesodium and 1 g magnesium stearate.

Purity analysis was conducted by reverse-phase HPLC (high performanceliquid chromatography). Impurities related to tenofovir disoproxilfumarate and emtricitabine were characterized and measured in the bulkAPI (active pharmaceutical ingredient) before formulation in the threelots of Table 2, and again after formulation in the resulting tablets.The impurities include by-products from hydrolysis of the exocyclicamino groups of tenofovir disoproxil fumarate and emtricitabine, and thehydrolysis of the disoproxil (POC) esters of tenofovir disoproxilfumarate. In each lot, the sum total of impurities related to tenofovirdisoproxil fumarate and emtricitabine was less than 1% after formulationand tablet manufacture.

The physicochemical properties of tenofovir disoproxil fumarate andemtricitabine tablets were evaluated by visual appearance, watercontent, label strength, impurity and degradation product contents, andtablet dissolution. Stability studies were conducted on drug productpackaged in container-closure systems that are identical to the intendedclinical and commercial container-closure system. There was no sign ofdiscoloration or tablet cracking during the course of the stabilitystudy. Film-coated tenofovir disoproxil fumarate and emtricitabinetablets exhibited satisfactory stability at 40° C./75% RH (relativehumidity) for up to six months when packaged and stored with silica geldesiccant. No significant loss (defined as >5% degradation) in % labelstrength of tenofovir DF or emtricitabine was observed after six monthsat 40° C./75% RH, when packaged and stored with desiccant. The increasein the total degradation products was 1.5% for tenofovir DF and 0.6-0.7%for emtricitabine after six months at 40° C./75% RH when packaged andstored with 3 grains of desiccant.

All publications and patent applications cited herein are incorporatedby reference to the same extent as if each individual publication orpatent application was specifically and individually indicated to beincorporated by reference.

Although certain embodiments are described in detail above, those havingordinary skill in the art will clearly understand that manymodifications are possible in the claims without departing from theteachings thereof. All such modifications are intended to be encompassedwithin the claims of the invention.

EMBODIMENTS OF THE INVENTION

A1. A pharmaceutical composition comprising an effective amount of acompound of the formula:

wherein R¹ and R² are independently selected from H, C₁-C₆ alkyl, C₁-C₆substituted alkyl, C₆-C₂₀ aryl, C₆-C₂₀ substituted aryl, C₆-C₂₀arylalkyl, C₆-C₂₀ substituted arylalkyl, acyloxymethyl esters—CH₂C(═O)R⁹ and acyloxymethyl carbonates —CH₂C(═O)OR⁹ where R⁹ is C₁-C₆alkyl, C₁-C₆ substituted alkyl, C₆-C₂₀ aryl and C₆-C₂₀ substituted aryl;

R³ is selected from H, C₁-C₆ alkyl, C₁-C₆ substituted alkyl, or CH₂OR⁸where R⁸ is C₁-C₆ alkyl, C₁-C₆ hydroxyalkyl and C₁-C₆ haloalkyl;

R⁴ and R⁵ are independently selected from H, NH₂, NHR and NR₂ where R isC₁-C₆ alkyl; and

R⁶ and R⁷ are independently selected from H and C₁-C₆ alkyl;

or a physiologically functional derivative thereof;

in combination with an effective amount of a compound of the formula

wherein B is selected from adenine, guanine, cytosine, uracil, thymine,7-deazaadenine, 7-deazaguanine, 7-deaza-8-azaguanine,7-deaza-8-azaadenine, inosine, nebularine, nitropyrrole, nitroindole,2-aminopurine, 2-amino-6-chloropurine, 2,6-diaminopurine, hypoxanthine,pseudouridine, 5-fluorocytosine, 5-chlorocytosine, 5-bromocytosine,5-iodocytosine, pseudocytosine, pseudoisocytosine, 5-propynylcytosine,isocytosine, isoguanine, 7-deazaguanine, 2-thiopyrimidine,6-thioguanine, 4-thiothymine, 4-thiouracil, O⁶-methylguanine,N⁶-methyladenine, O⁴-methylthymine, 5,6-dihydrothymine,5,6-dihydrouracil, 4-methylindole, and a pyrazolo[3,4-D]pyrimidine; and

R is selected from H, C₁-C₁₈ alkyl, C₁-C₁₈ substituted alkyl, C₂-C₁₈alkenyl, C₂-C₁₈ substituted alkenyl, C₁-C₁₈ alkynyl, C₂-C₁₈ substitutedalkynyl, C₆-C₂₀ aryl, C₆-C₂₀ substituted aryl, C₂-C₂₀ heterocycle,C₂-C₂₀ substituted heterocycle, phosphonate, phosphophosphonate,diphosphophosphonate, phosphate, diphosphate, triphosphate,polyethyleneoxy or a physiologically functional derivative thereof; and

a pharmaceutically acceptable carrier.

B2. A composition of embodiment A1 wherein, in formula 1, R¹ and R² areindependently selected from H, C₁-C₆ alkyl, C₁-C₆ substituted alkyl,C₆-C₂₀ aryl, C₆-C₂₀ substituted aryl, C₆-C₂₀ arylalkyl, C₆-C₂₀substituted arylalkyl, acyloxymethyl esters —CH₂C(═O)R⁹ andacyloxymethyl carbonates —CH₂C(═O)OR⁹ where R⁹ is C₁-C₆ alkyl, C₁-C₆substituted alkyl, C₆-C₂₀ aryl and C₆-C₂₀ substituted aryl; and R³R⁴,R⁵, R⁶ and R⁷ are independently H or C₁-C₆ alkyl.C3. A composition of embodiment A1 wherein, in formula 2, B is cytosineor a 5-halocytosine.D4. A composition of embodiment A1 wherein, in formula 1, R¹ and R² areindependently selected from H, C₁-C₆ alkyl, C₁-C₆ substituted alkyl,C₆-C₂₀ aryl, C₆-C₂₀ substituted aryl, C₆-C₂₀ arylalkyl, C₆-C₂₀substituted arylalkyl, acyloxymethyl esters —CH₂C(═O)R⁹ andacyloxymethyl carbonates CH₂C(═O)OR⁹ where R⁹ is C₁-C₆ alkyl, C₁-C₆substituted alkyl, C₆-C₂₀ aryl and C₆-C₂₀ substituted aryl; and R³ R⁴,R⁵, R⁶ and R⁷ are independently H or C₁-C₆ alkyl; and, in formula 2, Bis cytosine or a 5-halocytosine.E5. A composition of embodiment D 4 wherein, in formula 1, R¹ and R² areindependently selected from H, acyloxymethyl esters —CH₂C(═O)R⁹ andacyloxymethyl carbonates CH₂C(═O)OR⁹ where R⁹ is C₁-C₆ alkyl; and R³,R⁴, R⁵, R⁶ and R⁷ are independently H or C₁-C₆ alkyl; and, in formula 2,B is cytosine or a 5-halocytosine and R is H.F6. A composition of embodiment E5 wherein, in formula 1, R¹ and R² areindependently selected from H and —CH₂C(═O)OCH(CH₃)₂; R³ is —CH₃; andR⁴, R⁵, R⁶ and R⁷ are H; and, in formula 2, B is 5-fluorocytosine and Ris H.G7. A pharmaceutical composition comprising a pharmaceutically effectiveamount of [2-(6-amino-purin-9-yl)-1-methyl-ethoxymethyl]-phosphonic aciddiisopropoxycarbonyloxymethyl ester fumarate (tenofovir disoproxilfumarate) or a physiologically functional derivative thereof and apharmaceutically effective amount of(2R,5S)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-pyrimidin-2-one(emtricitabine) or a physiologically functional derivative thereof; anda pharmaceutically acceptable carrier.H8. A pharmaceutical formulation of embodiment A1 to G7 furthercomprising a third active ingredient selected from the group consistingof a protease inhibitor, a nucleoside or nucleotide reversetranscriptase inhibitor, a non-nucleoside reverse transcriptaseinhibitor, and an integrase inhibitor.I9. A pharmaceutical formulation of embodiments A1 to H8 in unit dosageform.J10. A method for the treatment or prevention of the symptoms or effectsof an HIV infection in an infected animal which comprises administeringto said animal a pharmaceutical composition of embodiments claims A1 to19.

We claim:
 1. A chemically stable fixed-dose combination comprising 300 mg of tenofovir disoproxil fumarate and 200 mg of emtricitabine wherein the combination exhibits less than 10% degradation of the tenofovir disoproxil fumarate and emtricitabine after six months at 40° C./75% relative humidity when packaged and stored with silica gel desiccant at 40° C./70% relative humidity.
 2. The chemically stable combination of claim 1 in the form of a pharmaceutical dosage form.
 3. The chemically stable combination of claim 2 wherein the dosage form is oral.
 4. The pharmaceutical dosage form of claim 2 wherein the tenofovir disoproxil fumarate is not substantially degraded.
 5. The pharmaceutical dosage form of claim 4 where there is less than 10% degradation of tenofovir disoproxil fumarate over a 24-hour period.
 6. The pharmaceutical dosage form of claim 4 where there is less than 1% degradation of tenofovir disoproxil fumarate over a 24-hour period.
 7. The pharmaceutical dosage form of claim 4 where there is less than 0.1% degradation of tenofovir disoproxil fumarate over a 24-hour period.
 8. The pharmaceutical dosage form of claim 4 where there is less than 0.01% degradation of tenofovir disoproxil fumarate over a 24-hour period.
 9. The pharmaceutical dosage form of claim 2 wherein less than 5% degradation of the tenofovir disoproxil fumarate and emtricitabine occurs after six months.
 10. A chemically stable fixed-dose combination comprising 300 mg of tenofovir disoproxil fumarate and 200 mg of emtricitabine wherein the combination exhibits less than 10% degradation of tenofovir disoproxil fumarate over a 24-hour period.
 11. The chemically stable combination of claim 10, in the form of a pharmaceutical dosage form.
 12. The chemically stable combination of claim 11, wherein the dosage form is oral.
 13. The pharmaceutical dosage form of claim 11, wherein there is less than 1% degradation of tenofovir disoproxil fumarate.
 14. The pharmaceutical dosage form of claim 11, wherein there is less than 0.1% degradation of tenofovir disoproxil fumarate.
 15. The pharmaceutical dosage form of claim 11, wherein there is less than 0.01% degradation of tenofovir disoproxil fumarate.
 16. The pharmaceutical dosage form of claim 11, wherein the combination exhibits less than 10% degradation of the tenofovir disoproxil fumarate and emtricitabine after six months at 40° C./75% relative humidity when packaged and stored with silica gel desiccant at 40° C./70% relative humidity.
 17. The pharmaceutical dosage form of claim 11 wherein the combination exhibits less than 5% degradation of the tenofovir disoproxil fumarate and emtricitabine after six months at 40° C./75% relative humidity when packaged and stored with silica gel desiccant at 40° C./70% relative humidity.
 18. The pharmaceutical dosage form of claim 2 or 11 comprising 300 mg tenofovir disoproxil fumarate, 200 mg emtricitabine, pregelatinized starch, croscarmellose sodium, lactose monohydrate, microcrystalline cellulose, and magnesium stearate.
 19. The pharmaceutical dosage form of claim 18 comprising 300 mg tenofovir disoproxil fumarate, 200 mg emtricitabine, 50 mg pregelatinized starch, 60 mg croscarmellose sodium, 80 mg lactose monohydrate, 300 mg microcrystalline cellulose, and 10 mg magnesium stearate.
 20. The pharmaceutical dosage form of claim 18 comprising 300 mg tenofovir disoproxil fumarate, 200 mg emtricitabine, 50 mg pregelatinized starch, 60 mg croscarmellose sodium, 180 mg lactose monohydrate, 200 mg microcrystalline cellulose, and 10 mg magnesium stearate.
 21. The pharmaceutical dosage form of claim 2 or 11 comprising 300 mg tenofovir disoproxil fumarate, 200 mg emtricitabine, pregelatinized starch, croscarmellose sodium, lactose monohydrate, microcrystalline cellulose, magnesium stearate, and colloidal silicon dioxide.
 22. The pharmaceutical dosage form of claim 21 comprising 300 mg tenofovir disoproxil fumarate, 200 mg emtricitabine, 50 mg pregelatinized starch, 60 mg croscarmellose sodium, 175 mg lactose monohydrate, 200 mg microcrystalline cellulose, 10 mg magnesium stearate, and 5 mg colloidal silicon dioxide.
 23. The pharmaceutical dosage form of claim 21 comprising 300 mg tenofovir disoproxil fumarate, 200 mg emtricitabine, hydroxypropyl methylcellulose, lactose B.P., pregelatinized starch B.P, and magnesium stearate.
 24. The pharmaceutical dosage form of claim 21 comprising 300 mg tenofovir disoproxil fumarate, 200 mg emtricitabine, 112 g hydroxypropyl methylcellulose, 53 mg lactose B.P., 28 mg pregelatinized starch B.P, and 7 mg magnesium stearate.
 25. The pharmaceutical dosage form of claim 2 or 11 comprising less than 1% of impurities related to tenofovir disoproxil fumarate and emtricitabine.
 26. The pharmaceutical dosage form of claim 2 or 11, further comprising a third anti-viral agent.
 27. The pharmaceutical dosage form of claim 26, wherein the third antiviral agent is selected from the group consisting of protease inhibitors, nucleoside reverse transcriptase inhibitors, non-nucleoside reverse transcriptase inhibitors, and integrase inhibitors.
 28. The pharmaceutical dosage form of claim 27, wherein the third antiviral agent is a protease inhibitor.
 29. The pharmaceutical dosage form of claim 27, wherein the third antiviral agent is a nucleoside reverse transcriptase inhibitor.
 30. The pharmaceutical dosage form of claim 27, wherein the third antiviral agent is a non-nucleoside reverse transcriptase inhibitor.
 31. The pharmaceutical dosage form of claim 27, wherein the third antiviral agent is an integrase inhibitor.
 32. The pharmaceutical dosage form of claim 30, wherein the third antiviral agent is efavirenz.
 33. A method for the treatment of the symptoms or effects of an HIV infection in an infected animal which comprises administering to said animal the pharmaceutical dosage form of claim 2 or
 11. 34. A method for the treatment of the symptoms or effects of an HIV infection in an infected animal which comprises administering to said animal the pharmaceutical dosage form of claim 9 or
 13. 35. A method for the treatment of the symptoms or effects of an HIV infection in an infected animal which comprises administering to said animal the pharmaceutical dosage form of claim
 21. 36. A method for the treatment of the symptoms or effects of an HIV infection in an infected animal which comprises administering to said animal the pharmaceutical dosage form of claim
 26. 37. A method for the treatment of the symptoms or effects of an HIV infection in an infected animal which comprises administering to said animal the pharmaceutical dosage form of claim
 27. 38. A method for the treatment of the symptoms or effects of an HIV infection in an infected animal which comprises administering to said animal the pharmaceutical dosage form of claim
 32. 